1. PRACTICAL GUIDE THE PROCESS OF TESTING
FOR USING THE NG DetecTool DEVICE

2. The advantages of the NG DetecTool
enables efficient patient treatment
prohibits further spread of ESBL and carbapenemase- producing bacteria
rapid and easy-to-use diagnostic system to detect ESBL and carbapenemases
in clinical samples:
direct detection of bacterial enzymes
responsible for ß-lactam resistance 2

3. Slow isolation methods
Current methods take hours and require cultured bacteria 3

4. The rapid immunoassay test
New detection system: the workflow for detection of multidrug resistant bacteria is shortened to min.
Workflow with NG DetecTool
Sample reception and processing
MDR bacteria detection / characterization
URINE
No required incubation
BLOOD CULTURE
2 min
RECTAL SWAB
overnight incubation
NGDETECTOOL 15 MINUTES 4

5. Sample types, what they show and next steps
RECTAL SWAB
isolation of the patient
COLONIZATION
BLOOD
treatment of the patient
INFECTION
URINE
Samples taken from blood cultures positive with gram-negative bacteria
Samples positive for gram-negative bacteria
Mintavételi eszközök képeknek 5

6. Key features of the NG DetecTool
Concentrates bacteria from liquid sample
Eliminates the interferences due to the biological media
Improved sensitivity
Decreases the time to result to minutes. Please note, incubation is required for rectal swab samples (overnight), which extends the time to result. 6

7. Detection process and features
One device Two functions
Sample processing
Filtration
Extraction
Incubation
Deposition
B. Beta-lactamases Detection
Lateral Flow immunoassay (LFIA) (Similar technology as a pregnancy test)
Single use
No specific equipment
ATTENTION: Please make sure you read the detailed NG DetecTool protocol before using the device on real-life samples. 7

8. Detection system components 1
Filtration part:
Filtration / Concentration
Upper part
Membrane 0,2µm
Cup
Lower part 8

9. Detection system components 2
Processing part:
Three main components
Cap/plunger
Membrane 0,2 µm
Cup
Tank 9

10. Detection system components 3
Detection part:
Two main components
Cassette
Strip
Test lines indicate the presence of enzymes responsible for the antibiotic resistance 10

11. Detection process – Sample preparation
This training material is a high-level overview on the operation of the device. ATTENTION: Please make sure you read the detailed NG DetecTool protocol before using the device on real-life samples.
BLOOD
URINE
Mix 2 volumes of positive blood culture with 1 volume of lysis buffer. Let react for minimum 2 minutes (possible up to 30’).
RECTAL SWAB
No preparation required
Inoculate transport medium of the swabs into Brain heart infusion media supplemented with cefotaxime for ESBL (CTXM) and ertapenem (for carbapenemases) and incubate overnight.
Mintavételi eszközök képeknek 11

12. Detection process – Filtration A
Sampling and filtration
Sample volume ≤ 1,5 ml
1) Insert the cup into
the lower part
2) Add the sample
in the cup
3) Close the filtration system
with the upper part
4) Push the sample through the filter using a syringe full of air
Sample
Bacteria 12

13. Detection process – Filtration B
Sampling and filtration
Sample volume ≥ 1,5 ml
1) Insert the cup into
the lower part
2) Close the filtration system
with the upper part
3) Aspirate the desired volume of sample and some air with a syringe
4) Push the sample and
the air through the filter
Bacteria 13

14. Detection process – Extraction
Extraction and incubation
1) Remove the lower part
2) Transfer the cup in the tank and remove the upper part
Bacteria
Labelled
antibodies 14

15. Detection process – Extraction
Extraction and incubation
1) Remove the lower part
2) Transfer the cup in the tank and remove the upper part
3) Add 250 μl extraction buffer in the cup
4) Screw the cap
5) Shake slightly the processing part (to conjugate the dissolution)
Extraction buffer
Labelled
antibodies 14

16. Detection process – Detection
1) Place the processing part on the detection part
Extracted sample +
Labelled antibodies
Strip 15

17. Detection process – Detection
1) Place the processing part on the detection part
2) Break the bottom of the processing part by pressing it down
3) Read the result after 15 minutes 15

18. Principle of the detection
Immunoassay:
antigene (analyte) – antibody reaction
Lateral Flow ImmunoAssay: the sample is applied at the end of the strip (sample pad), then migrates into the detection zone
Antibody
Enzyme (Antigen)
Labelled antibody
Sample
Migration direction
Test line
Control line
Sample pad
Detection zone 16

19. Detection process – Detection
The strip contains:
a control line to confirm that the test is working properly
testline(s) - ESBL(CTX-M) or carbapenemases (KPC, OXA, VIM, IMP, NDM)
Strip to ESBL
Strip to carbapenemases
Control line
KPC
OXA
CTX-M
VIM
IMP
NDM
Sample pad 17

20. Detection process – Evaluation
Read the result after 15 minutes
The control line must be visible: when the control line does not appear, the test is invalid
Absence of a testline -> NEGATIVE
Presence of a testline -> POSITIVE 18

21. When the result is positive The patient must be placed in isolation
Actions to be taken 1
When the result is positive
From rectal swab
From blood / urine
COLONISATION
INFECTION
The patient must be placed in isolation
Antibiotic treatment 19

22. Actions to be taken 2
Positive result: consultation with infection control specialist is necessary
Limited therapeutic options, consultation with infectious disease specialist
Antibiotic susceptibility test is required to adapt best therapy
Negative result for blood or urine sample:
the sample may contain other resistant bacteria (except for the ESBL- and carbapenemase-producing bacteria)
for haemoculture and urine, bacterial culture is required
Negative result for rectal swab:
no action required 20

23. This is the end of the Principles of Testing Section of the NG DetecTool training Thank you for your attention!